Two-dimensional protein crystals on a solid substrate: Effect of surface ligand concentration
Two-dimensional protein crystals on a solid substrate: Effect of surface ligand concentration
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DOI：
10.1021/la701399s
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发表时间：
2007-09-11
期刊：
影响因子：
3.9
通讯作者：
Wang, Szu-Wen
中科院分区：
文献类型：
--
作者：
Lou, Chengfei;Wang, Zifu;Wang, Szu-Wen
包埋在固体支持的脂质双层中的蛋白质可以作为研究细胞膜和蛋白质在表面上行为的模型系统。我们研究了链霉亲和素在云母支撑的双层膜上的自组装。使用荧光显微镜和原子力显微镜，我们的研究表明，表面配体的浓度影响所得蛋白质阵列的分子堆积，这反过来又影响整体晶体形态。当底物上的生物素化脂质密度达到1.5%摩尔分数时，获得二维链霉亲和素晶体，产生主要由具有P1对称性的晶体组成的高纵横比形态。在3%及以上时，形成具有C222对称性的晶体，并导致H形和汇合结构。在2和3%之间的中间密度下，观察到PI和C222晶型的共存。宏观形态和分子构型之间的关系是类似于以前报道的数据在空气/水界面处获得。这表明，在我们的实验条件下，蛋白质与支持底物的相互作用对于定义自组装行为的意义小于蛋白质分子之间的相互作用。配体抑制和光漂白后的荧光恢复被用来阐明浓度依赖性机制的分歧晶体形式。我们已经测量的扩散系数的分子在P1形成的条件下，大约是两倍的分子在C222形成的浓度，这是一致的蛋白质结合到表面通过一个和两个配体，分别。因此，与结合状态相关的差异灵活性可能会改变结晶中涉及的微妙蛋白质相互作用。
Proteins imbedded in solid-supported lipid bilayers can serve as model systems for investigations of cellular membranes and protein behavior on surfaces. We have investigated the self-assembly of streptavidin on mica-supported bilayer membranes. Using fluorescence microscopy and atomic force microscopy, our studies reveal that the concentration of surface ligand influences the molecular packing of the resulting protein arrays, which in turn affects overall crystal morphology. Two-dimensional streptavidin crystals are obtained when the biotinylated lipid density on the substrate reaches 1.5% mole fraction, yielding high-aspect morphologies that comprise primarily of crystals with P1 symmetry. At 3% and above, crystals with C222 symmetry are formed and result in H-shaped and confluent structures. In intermediate densities between 2 and 3%, a coexistence of PI and C222 crystal forms is observed. The relationship between macroscopic morphology and molecular configuration is similar to previously reported data obtained at the air/water interface. This suggests that, under our experimental conditions, protein interactions with the supporting substrate are less significant for defining self-assembly behavior than interactions between protein molecules. Ligand-inhibition and fluorescence recovery after photobleaching were used to elucidate the concentration-dependent mechanism for the divergent crystal forms. We have measured the diffusion coefficient of molecules in P1-forming conditions to be approximately twice that of molecules in C222-forming concentrations, which is consistent with proteins bound to the surface through one and two ligands, respectively. The differential flexibility associated with the binding state is therefore likely to alter the subtle protein interactions involved in crystallization.